AB0134 IN-VITRO STUDY ON THE EFFECT OF SELECTIVE Jak-INHIBITORS ON PBMCs STAT3 PHOSPHORYLATION FROM SYSTEMIC SCLEROSIS PATIENTS
Authors: F. Cacciapaglia, S. Perniola, S. del Vescovo, S. Stano, R. Bizzoca, D. Natuzzi, M. Fornaro, F. Iannone
Journal: Annals of the Rheumatic Diseases (2022)

Background Systemic sclerosis (SSc) is a rare autoimmune connective tissue disease characterized by autoimmunity-driven damage and vasculopathy leading to fibrosis of the skin and internal organs (1). The Janus kinase (Jak) - signal transducer and activator of transcription (STAT) pathway has been evidenced markedly activated in SSc patients (2, 3), and its inhibition has been proved in preclinical and clinical trials (4), but no data on Jak selective inhibition are available. Objectives To explore the effect of selective inhibition of Jak/STAT pathway in peripheral blood mononuclear cells (PBMC) from SSc patients. Methods In vitro Jak inhibition of the subunit 3 of phosphorylated (p) than activated STAT was measured by flow cytometry in peripheral blood mononuclear cells (PBMC) from SSc patients naïve to any immunosuppressive and/or corticosteroids (n.5). pSTAT3 activity was also assessed after stimulation with recombinant human 0.1 ng/ml IL-6 (Peprotech – NJ, USA). The PBMC were overnight incubated with IC50 concentrations of selective Jak1-, Jak2-, Jak3- and Tyk2-inhibitors (Biovision Inc. – CA, USA). Percentages of pSTAT3 positive cells were compared in presence of different compounds stimulation. Results After overnight incubation, percentage of pSTAT3 positive cells was significantly higher in CD14 pos compared to CD4 pos (16.3%; 95CI 10-22 vs 10.7%; 95CI 4--18, – p=0.02). pSTAT3 pos CD14 pos cells were halved only by selective Jak3-inhibitor, while pSTAT3 pos CD4 pos cells were reduced by 36% by selective Jak1-inhibitor. Selective Jak2- or Tyk2-inhibitors did not interfere with STAT3 phosphorylation in PBMC from SSc patients. After IL-6 stimulation, we observed a 2- and a 1.5-fold increase in percentage of pSTAT3 pos CD4 pos and pSTAT3 pos CD14 pos cells, respectively. pSTAT3 pos CD14 pos cells were reduced in the PBMC co-culture with IL-6 and Jak-selective inhibitors, in contrast no effects were found in CD4 pos cells. Specifically, selective Jak1- and Jak3-inhibitors reduced pSTAT3 pos CD14 pos cells by an average of 37% and 25%, respectively. No effects were observed after co-culture with IL-6 and selective Jak2- or Tyk2-inhibitors. Conclusion Jak/STAT3 pathway of PBMC from SSc patients with active disease may be differently modulated by specific inhibitors. Selectivity of Jak1- and Jak3-inhibitors seems more relevant, especially in CD14 pos monocytes after IL-6 stimulation. These preliminary findings highlight some evidence for effectiveness of selective Jak-inhibitors in SSc treatment. References [1]Benfaremo D, et al. Systemic Sclerosis: From Pathophysiology to Novel Therapeutic Approaches. Biomedicines. 2022;10(1):163. [2]Talotta R. The rationale for targeting the JAK/STAT pathway in scleroderma-associated interstitial lung disease. Immunotherapy. 2021;13(3):241-256. [3]Cacciapaglia F, et al. Phosphorylated signal transducer and activator of transcription 3 (pSTAT3) is highly expressed in CD14+ circulating cells of scleroderma patients. Rheumatology (Oxford). 2020;59(6):1442-1444. [4]Karalilova RV, et al. Tofacitinib in the treatment of skin and musculoskeletal involvement in patients with systemic sclerosis, evaluated by ultrasound. Rheumatol Int. 2021;41(10):1743-1753. Disclosure of Interests None declared